In-vitro cell dynamic culture device with independent unit design

Abstract

The invention discloses a device suitable for high-throughput detection of biological effects of materials, the device can be used independently, does not need to be placed in an incubator, and simulates in-vivo microenvironment culture in an in-vitro cell dynamic and circulating mode. The device comprises a culture chamber, a liquid changing chamber, a culture environment maintaining system, a constant temperature system, a power system and the like, the culture chamber adopts a high-throughput screening reaction chamber with independent unit design, mutual interference among different groups can be avoided, and high-throughput screening of different physical and chemical properties such as structures and components of biological materials can be realized; the efficiency is improved; and parallel operation of experiments is ensured. The culture environment maintaining system and the constant-temperature system ensure a good environment in the cell culture process; the power system is used for simulating an in-vivo circulation system, so that the culture medium is ensured to be in a flowing and real-time updated culture environment, and experimental errors caused by limitation of a narrow space and a non-circulation state in a biological material research and development process are avoided.

Description

Technical Field

[0001] The invention belongs to the field of biomedical materials, and in particular relates to an independent device for in vitro dynamic cell culture, which is suitable for designing a high-throughput screening device for the physical and chemical properties of biological materials. Background Art

[0002] Cell culture refers to a method of simulating the in vivo environment in vitro to enable it to survive, grow, reproduce and maintain its main structure and function. Cell culture is also called cell cloning technology, and its formal term in biology is cell culture technology. Cell culture is an indispensable process for both the entire bioengineering technology and one of its biological cloning technologies. Cell culture technology is an important and commonly used technology in cell biology research methods. Through cell culture, a large number of cells can be obtained, and cell signal transduction, cell anabolism, cell growth and proliferation, etc. can be studied. It is of great significance to the development of drugs, biopharmaceuticals, and biomaterials.

[0003] However, the in vitro culture model currently used is static and space-limited and cannot simulate the in vivo environment. When used in the research and development of drugs and biomaterials, the limited static non-circulating in vitro culture space often leads to experimental errors. For example, in the study of biomaterial components, the limited static non-circulating in vitro culture space can lead to excessive local element concentrations or rapid changes in pH due to dissolution and degradation, resulting in cytotoxicity. In addition, the study of sustained-release of drug-loaded materials will also be subject to differences due to the limited static non-circulating in vitro culture space. The dynamic cyclic in vitro culture system used in this invention is suitable for the research and development of related materials and drugs, and adopts a high-throughput screening culture unit integration model. Each culture unit is independently designed, which is conducive to the simultaneous biological detection of multiple samples, improves efficiency, and can avoid differences caused by repeated experiments due to human errors, thereby improving the reliability of experimental data.

[0004] A representative example is the technical patent High-throughput 3D cell, tissue-like and organoid dynamic culture system (CN202310325355.7), which describes a high-throughput 3D cell, tissue-like and organoid dynamic culture system, including a casing body, a culture solution mixing tank arranged in the casing body in sequence along the culture solution delivery direction, a peristaltic pump, a perfusion culture chamber, a waste liquid storage chamber, and a pure gas source arranged on the side of the casing body and connected to the culture solution mixing tank. However, in the above patents, the high-throughput chip does not adopt an independent design, and has limitations in the research and development of large-size scaffold materials. In addition, closed hoses are used in series between the various parts, and a filter cover design is used in the fluid exchange chamber and the culture chamber to ensure the balance of air pressure in each area, which is more convenient and sterile to realize routine experimental operations such as culture medium replacement and sample observation, and is more in line with the operation under the existing cell culture technical specification experimental process. Summary of the invention

[0005] In order to better simulate the dynamic and cyclic processes in the in vivo culture environment and reduce the experimental errors caused by the limited static non-cyclic in vitro culture space in the research and development of drugs and biomaterials, the present invention discloses a device suitable for high-throughput screening of the physical and chemical properties of biomaterials, which simulates the in vivo microenvironment culture by using in vitro cell dynamics and cycles.

[0006] In view of the above problems, the object of the present invention is achieved by at least one of the following solutions:

[0007] A device suitable for high-throughput screening of biomaterials or the design of physical and chemical properties of biomaterials, which simulates in vivo microenvironment culture by in vitro cell dynamics and circulation. The device includes a culture chamber, a fluid exchange chamber, a culture environment maintenance system, a constant temperature system and a power system, etc. The various parts are connected by closed hoses. At the same time, the fluid exchange chamber and the culture chamber are designed with filter covers to ensure air pressure balance in each area. BRIEF DESCRIPTION OF THE DRAWINGS

[0008] Figure 1 Schematic diagram of the overall device

[0009] Figure 2 It is a high-throughput screening reaction chamber with independent culture chamber and independent unit design, which can be arranged in parallel.

[0010] Figure 3 To maintain the atmosphere, temperature and other environmental conditions during cell culture

[0011] Specific implementation

[0012] The specific examples listed are only used to further illustrate the present invention, but the implementation of the present invention is not limited to the real-time examples listed.

[0013] Example 1

[0014] High-throughput screening of different physical and chemical properties of biomaterials such as structure and components.

[0015] Devices such as Figure 1 As shown, it includes a culture chamber, a fluid exchange chamber, a culture environment maintenance system, a constant temperature system, and a power system. Different samples to be tested are placed in the well plate of the culture chamber. The culture chamber is an independent unit, and different groups will not affect each other. The culture medium is injected into the fluid exchange chamber, and the culture environment maintenance system is adjusted as shown in FIG. Figure 3 As shown, the environment maintenance system uses a carbon dioxide cylinder to deliver carbon dioxide into the culture device to maintain the carbon dioxide concentration in the culture environment, and the air inlet can bring a certain temperature and a moderate atmosphere into the culture device in warm water to maintain the culture environment of the device. The environment maintenance system connects the culture room and the fluid exchange room. After the environment required for cell culture (37°C, 5% carbon dioxide) is reached in the device, the cell plate is plated. This part of the experimental operation is carried out in a sterile environment. Different samples to be tested can be biomaterials with different components, tissue engineering scaffolds with different structures, etc., to compare the mediating effects of different types and materials after treatment processes on cells.

[0016] After the cells are plated, the device does not need to be planted in a sterile environment. Turn on the power system to add the fresh culture medium in the fluid exchange chamber to the culture chamber. The power system uses a peristaltic pump, and the speed of the peristaltic pump can be set. The outlet position in the culture chamber is close to the bottom position to ensure that the fresh culture medium enters from the top during the flow of the culture medium, and the old culture medium circulates out of the culture chamber from the bottom and flows into the fluid exchange chamber. The fresh culture medium and the old culture medium in the fluid exchange chamber are separated and stored independently, and the old culture medium is stored in the sampling and detection chamber, and the relevant secretory products can be sampled. The device as a whole is a semi-closed setting. Except for the culture chamber and the fluid exchange chamber, most of the other devices are closed designs. The outer shell of the culture chamber and the fluid exchange chamber is installed with a semi-permeable membrane with antibacterial properties to maintain the exchange and pressure of the external atmosphere.

[0017] Example 2

[0018] Reveal the mechanism of action of biological and basic medical processes, and study the interaction between cells, tissues, organs and paracrine mechanisms

[0019] The culture chamber in this device is an independent unit. The outlet position of the orifice plate of different culture chambers can be connected in parallel with the inlet position of the next orifice plate to form a series of connected culture chambers. The environmental culture medium in the first culture well is added to the next culture well by using the principle of U-shaped tube. This setting can not only be used for the setting of parallel samples of experimental groups, but also for studying the interaction and paracrine mechanism between different cells, tissues and organs. The culture medium is injected into the liquid exchange chamber, and the culture environment maintenance system is adjusted. Figure 3As shown, the cell culture is carried out after the environment required for cell culture (37°C, 5% carbon dioxide) is reached in the device. For example, bone marrow mesenchymal stem cells are added to culture well 1 in the culture chamber, and endothelial cells are added to culture well 2. The effect of the paracrine effect of bone marrow mesenchymal stem cells on endothelial cell vascularization can be studied, and the osteogenic / angiogenic coupling effect can be studied. The interaction and paracrine effect between related tissues and cells can also be achieved through this device.

[0020] After the cells are plated, the power system is turned on to add the fresh culture medium in the fluid exchange chamber to the culture chamber, ensuring that the fresh culture medium enters from the top during the flow of the culture medium, and the old culture medium circulates out of the culture chamber from the bottom and flows into the fluid exchange chamber. The fresh culture medium and the old culture medium in the fluid exchange chamber are separated and stored independently, and the old culture medium is stored in the sampling and detection chamber, and the relevant secretory products in it can be sampled and analyzed.

Claims

1. A device suitable for high-throughput screening of physical and chemical properties of biological materials, the device comprising a culture chamber, a liquid exchange chamber, a culture environment maintenance system, a constant temperature system, and a power system, etc.: (1) Culture environment maintenance system and constant temperature system: The device can be used independently and does not need to be placed in a cell culture box; (2) The in vitro cell culture process adopts a dynamic and cyclical approach; (3) The culture room adopts a high-throughput screening culture unit integrated model, and each culture unit adopts an independent design.

2. The constant temperature system according to claim 1, characterized in that: The temperature of the culture chamber and the fluid exchange chamber can be maintained at a temperature suitable for cell culture, preferably 37°C.

3. The constant temperature system for the culture environment maintenance system according to claim 1, characterized in that Provides a mild, environmental atmosphere suitable for cell growth in the culture chamber, as well as auxiliary temperature control.

4. The dynamic, cyclical culture method according to claim 1, characterized in that: A power system such as a peristaltic pump is used to bring fresh culture medium from the fluid exchange chamber into the culture chamber, while the old culture medium in the culture chamber is sucked out to maintain freshness.

5. According to claim 1, the culture chamber adopts a linked tube mode, including specifications such as six-well plates and twenty-four-well plates, to meet the needs of high-throughput screening of the physical and chemical properties of biomaterials, and each independent culture chamber has an independent culture medium inlet and liquid exchange outlet channel to avoid interference between samples of different groups.

6. The device for high-throughput screening of physical and chemical properties of biomaterials according to claim 2, characterized in that: The well plate in the culture chamber and the relative medium exchange channel can be adjusted according to the number of samples, ranging from 1 to 100, while the medium outlet is kept below the highest liquid level.

7. The independent device for in vitro dynamic cell culture according to claim 3, characterized in that: Each part is connected with a closed rubber hose, and the liquid exchange room and culture room are designed with filter covers to ensure the air pressure balance in each area.

8. The independent device for in vitro dynamic cell culture according to claim 4, characterized in that: Step (1) The liquid exchange chamber is partitioned to separate the fresh culture medium from the old culture medium. The fresh culture medium area is designed with a detachable filter cover to facilitate the addition of culture medium.

9. The independent device for in vitro dynamic cell culture according to claim 5, characterized in that: Cycle time, power speed adjustment, and includes a low temperature alarm system, using low temperature exploration for detection.

10. An independent device for in vitro dynamic cell culture according to any one of claims 1 to 9, characterized in that: Can maintain long-term, independent cell cultures.

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